AGP31 Antibody
Description
AGP31 Protein Overview
AGP31 (encoded by At1g28290) is a chimeric cell-wall glycoprotein characterized by:
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A multi-domain structure: Signal peptide, His-rich region, Pro-rich domain, and a C-terminal PAC domain .
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Glycosylation: Comprises ~80% galactose and ~13% arabinose, with Hyp-O-glycosylation on its Pro-rich domain .
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Localization: Primarily in vascular tissues and reproductive organs (e.g., embryo sacs) .
Antibody Applications in AGP31 Research
Key studies utilizing antibodies for AGP31 characterization include:
Western Blot Detection
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Anti-myc antibody: Used to detect AGP31-myc fusion proteins, confirming AGP31’s migration as a high-molecular-mass (170–200 kDa) glycoprotein due to extensive glycosylation .
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Anti-V5 antibody: Identified interactions between AGP31’s PAC domain and cell-wall polysaccharides (e.g., galactan, rhamnogalacturonan I) .
Immunolocalization
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Yariv reagent: A β-glucosyl synthetic phenylglycoside used to precipitate AGP31, confirming its classification as an AGP .
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Lectin probes: Peanut agglutinin (PNA) binding demonstrated AGP31’s galactan-rich glycosylation .
Glycosylation Analysis
| Parameter | Data | Source |
|---|---|---|
| Monosaccharides | 80.9% Gal, 13.5% Ara, 2.1% Xyl | |
| Amino Acids | Pro (14.8%), Hyp (14.4%), His (10.5%) | |
| Molecular Mass | 50–90 kDa (SDS-PAGE) vs. 38 kDa (predicted) |
Functional Insights
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Cell-wall interactions: AGP31 binds galactan and rhamnogalacturonan I via its PAC domain .
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Developmental roles: Expressed in vascular bundles and reproductive tissues, suggesting roles in cell-wall reinforcement during growth .
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Stress response: mRNA levels decrease under methyl jasmonate (MeJA) or wounding, mediated by COI1 signaling .
Technical Challenges and Solutions
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Glycosylation heterogeneity: AGP31 exists as multiple glycoforms, requiring deglycosylation (e.g., TFMS treatment) for accurate MS analysis .
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Antibody specificity: Recombinant tags (e.g., myc, V5) mitigate cross-reactivity with endogenous proteins .
Future Directions
Product Specs
**Constituents:** 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- AGP31 interacts with galactans, which are branches of rhamnogalacturonan I, through its PAC domain. PMID: 24685714
- The Pro-rich domain in AGP31 is predominantly glycosylated. PMID: 22270363
- AGP31 may play a role in vascular tissue function during both defense responses and developmental processes. PMID: 17885091
Q&A
Arabinogalactan protein 31 (AGP31) antibodies are critical tools for studying plant cell wall dynamics and protein-polysaccharide interactions. Below are structured FAQs addressing key research considerations, organized by complexity and supported by experimental evidence from peer-reviewed studies.
Advanced Research Questions
How to investigate AGP31's role in rhamnogalacturonan I (RG-I) scaffolding?
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Stepwise protocol:
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Isolate RG-I from Arabidopsis hypocotyls via size-exclusion chromatography ( ).
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Perform solid-phase binding assays with nitrocellulose-spotted RG-I and AGP31 (10-100 µg/mL).
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Quantify PAC domain contribution using truncated mutants (e.g., ΔPAC-AGP31) ( ).
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Validate in planta via FRET-FLIM with RG-I-specific antibodies ( ).
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Does AGP31 function as a boron-dependent RG-II chaperone?
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Key findings:
How to resolve contradictions in AGP31's molecular weight reports?
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Analysis framework:
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Compare deglycosylation effects: Treat with TFMS (anhydrous trifluoromethanesulfonic acid) to remove arabinogalactans. Post-treatment MW shifts from 50-90 kDa → ~38 kDa confirm glycan contribution ( ).
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Correlate developmental stage with glycoform distribution: Etiolated hypocotyls show 90 kDa dominance vs. 50 kDa in mature tissues ( ).
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Technical Optimization
Strategies for improving AGP31 antibody specificity in IHC
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Protocol enhancements:
Quantitative profiling of AGP31 isoforms
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Workflow:
Emerging Research Frontiers
Engineering AGP31 variants for enhanced RG-II crosslinking
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Rational design:
AGP31 orthologs in crop species: Conservation and divergence
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Comparative analysis:
